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Several aldehyde dehydrogenase (ALDH) complexes have been purified from the membranes of acetic acid bacteria. The enzyme structures and the chemical nature of the prosthetic groups associated with these enzymes remain a matter of debate. We report here on the molecular and catalytic properties of the membrane-bound ALDH complex of the diazotrophic bacterium Gluconacetobacter diazotrophicus. The purified ALDH complex is a heterodimer comprising two subunits of 79.7 and 50 kDa, respectively. Reversed-phase high-pressure liquid chromatography (HPLC) and electron paramagnetic resonance spectroscopy led us to demonstrate, for the first time, the unequivocal presence of a pyrroloquinoline quinone prosthetic group associated with an ALDH complex from acetic acid bacteria. In addition, heme b was detected by UV-visible light (UV-Vis) spectroscopy and confirmed by reversed-phase HPLC. The smaller subunit bears three cytochromes c. Aliphatic aldehydes, but not formaldehyde, were suitable substrates. Using ferricyanide as an electron acceptor, the enzyme showed an optimum pH of 3.5 that shifted to pH 7.0 when phenazine methosulfate plus 2,6-dichlorophenolindophenol were the electron acceptors. Acetaldehyde did not reduce measurable levels of the cytochrome b and c centers; however, the dithionite-reduced hemes were conveniently oxidized by ubiquinone-l; this finding suggests that cytochrome b and the cytochromes c constitute an intramolecular redox sequence that delivers electrons to the membrane ubiquinone. doi: 10.1128/JB.00589-10

A study investigates whether the near-IR fluorescent probe Nile Red is an allosteric substrate of cytochrome P450 3A4 (CYP3A4). It is concluded that Nile Red, which undergoes sequential oxidation and exhibits complex binding behavior, provides the advantage of extreme solvatochromism and can be used to explore the properties any sites on CYP3A4 to which it binds.

Two well-studied small alpha-helical proteins, the B-domain of protein A and Rd-apocytochrome b562 were coupled by fusing the C-terminal helix of the B-domain of protein A with the N-terminal helix of Rd-apocyt b562 without changing their hydrophobic core residues. The results suggest that the two domains have weakenergetic coupling.

The pH dependencies of redox difference spectra is analyzed to study the redox-linked protonation chemistry of the iron-sulfur protein (ISP) of the cytochrome bc1 complex by using perfusion/electrochemically induced attenuated total reflection-Fourier transform infrared (ATR-FTIR) spectroscopy. Histidine is relatively sharp and has a high extinction coefficient and also, provides a novel marker band for the detection of imidazolate intermediates in enzymatic mechanisms.

X-ray structures at 3.0-3.1 angstrom resolution of the cytochrome b6f complex from the cyanobacterium Mastigocladus laminosus and the green alga Chlamydomonas reinhardtii showed the presence of a unique heme, heme x. The only axial ligand associated with this heme is a H2O or OH(super -) that is H-bonded to the propionate of the stormal side heme bn, showing that it is pentacoordinate.

Pseudoazurin binds at a single site on cytochrome c peroxidase from Paracoccus pantotrophus with a K(sub d) of 16.4 MuM at 25 degree C, pH 6.0, in an endothermic rdcytochromeeaction that is driven by a large entropy change. Cytochrome c550 and pseudoazurin bind at the same site on the cytochrome c peroxidase and that the pair of electrons required restoring the enzyme to its active state after turnover are delivered one-by-one to the electron-transferring heme.

A detailed structural, spectroscopic and thermodynamic characterization of cytochrome P450 BioI is conducted and reported. Results demonstrate unexpected flexibility of the active site with the capacity to bind tightly to certain steroids and to bulky azole drugs, as well as long-chain fatty acids. The result also reveals that BioI is purified from Escherichia coli with palmitic acid bound, which explains its isolation in a mixed spin state.

The time-resolved reaction between fully reduced cytochrome [ba.sub.3] from Thermus thermophilus and [O.sub.2] in variants where Thr-312 and Thr-315 are modified is examined. The results have shown that proton uptake is inhibited in the two variants, which has suggested involvement of T312 and T315 in the transfer of protons to the achieve site during [O.sub.2] reduction in the [ba.sub.3] oxidase.

A study was conducted to understand the kinetic mechanism related to the role of structurally homologous flavoproteins photolyases and cryptochromes (CRY) towards UV-damaged DNA repair by photoinduced electron transfer from their FAD cofactor. The results obtained offer insight into the kinetic tuning of the rates of electron transfer which would play a function-defining role in photolyases and cryptochromes.

Cryogenic Fourier transform infrared (FTIR) photolysis experiments on fully reduced [bo.sub.3]-NO and the mixed gas [bo.sub.3]-CO/NO complexes are described. The difference in metal-metal distance has not affected the NO reductase activity and hence the results have shown that the coordination of the second NO to [Cu.sub.B] is not a necessary step of the reaction mechanism.

Microspectrophotometry of single crystals of cytochrome [ba.sub.3] oxidase from Thermus thermophilus has shown that crystals of the oxidized enzyme are reduced in an intense X-ray. An unexpected optical absorption envelope at approximately 590 nm is observed from a five-coordinate, low-spin, ferrous heme [a.sub.3] that is trapped in the frozen crystals.

The solution structure of the reduced form of Shewanella frigidimarina NCIMB400 (Sfc) is analyzed. The studies have shown that the small tetraheme cytochromes isolated from Shewanella spp. has represented a new structural motif that is distinct from that of the tetraheme cytochromes [c.sub.3] from the Desulfovibrionacea family.

A stopped-flow technique in conjunction with theoretical computation method is used to examine the kinetics of reduction of a series of Fe(III) complexes with citrate, nitrilotriacetic acid (NTA), and EDTA by MtrC and OmcA. The results revealed that ligand complexation and relatively large reorganization energy contributes significantly to microbial dissimilatory reduction and mineral transformation of iron, as well as other redox-sensitive metal species in nature.

The glutamic acid residue of transmembrane helix III in subunit I (E107L) mutant cytochrome bd from Escherichia coli is studied by using electrometry and absorption spectroscopy with a microsecond time resolution to elucidate the role of E107 in translocation of protons within the protein molecule. The studies have shown that E107 is either the second group or a key residue of a proton delivery pathway leading from the cytoplasm toward the second group.

The characterization and functions of the petJ1 and petJ2 cytochromes isolated from cyanobacterium Synechococcus PCC 7002 are described. The electrostatic and redox properties of the two cytochromes are found to differ radically.

The basic thermodynamic and kinetic parameters for the reaction of cytochrome bd from Azotobacter vinelandii with oxygen was measured and compared with those of the Escherichia coli enzyme. The results suggest that in cytochrome bd, the redox state of the b-type hemes (heme [b.595]), controls the pathway for ligand transfer between heme d and the bulk phase.

The reaction in solution is studied to understand how the very weak binding between the two proteins from Phormidium laminosum can nevertheless lead to fast rates of electron transfer. The results are interpreted in terms of a model in which the stable form of the protein-protein complex requires further thermal activation to be competent for electron transfer.

A single crystal of the bovine mitochondrial cytochrome bc1 complex with stigmatellin in the Q (super 0) quinol binding site determines the orientation of the g-tensors of the Rieske iron-sulfur protein subunit. Each principal g-tensor has important implications, for the electronic structure of the 2Fe2S cluster and its role in electron transfer reactions.

Research has been conducted on ester-containing 5-carboxyl-5-methyl-1-pyrroline N-oxides. The estimated rate constants for these oxides at acidic and neutral pH for the reaction of ester-containing 5-carboxyl-5-methyl-1-pyrroline N-oxides with superoxide have been achieved via the use of ferricytochrome c.